ε-聚赖氨酸对白念珠菌抑菌活性及机制研究

目的 研究ε-多聚赖氨酸(ε-poly-L-lysine, ε-PL)对白念珠菌(Candida albicans)的抑菌活性及抑菌机制。方法 以白念珠菌的标准菌株ATCC64548(氟康唑敏感株)、ATCC64550(氟康唑耐药株)以及临床收集的50株菌为实验菌株，按照CLISI- M27文件中的微量稀释法测定ε-多聚赖氨酸的MIC、MFC和SMIC50值；绘制48h内的浮游菌株时间-生长曲线和生物膜抑制-时间曲线；连续观测并记录4h内的芽管形成率和芽管长度；测定药物处理前后白念珠菌的丙二醛和活性氧(ROS)含量。结果 ε-PL对白念珠的最低抑菌浓度(MIC)为512μg/mL，最低杀菌浓度(MFC)为1024μg/mL，SMIC50为512μg/mL，ε-PL对白念珠菌浮游菌及生物膜的抑菌作用随着浓度的升高作用愈明显。抑菌-时间曲线结果表明ε-PL对白念珠菌的浮游菌和生物膜在12h左右即产生明显抑制作用。芽管实验的结果表明高浓度的ε-PL对白念珠菌的芽管形成率及芽管长度具有明显抑制作用。ε-PL作用于白念珠菌后MDA和ROS含量呈现上升趋势，并且与药物浓度大小成正相关。结论 ε-PL对白念珠菌浮游菌株及生物膜均有良好的抑制作用，高浓度ε-PL对白念珠菌主要毒力菌丝有明显抑制作用，ε-PL作用导致白念珠菌内产生大量的活性氧(ROS)以及产生一定程度的脂质氧化，提示ε-PL可能通过氧化作用发挥抑菌效果。

In vitro antifungal activity of ε-poly-lysine against Candida albicans

Abstract Objective To investigate the antifungal activity and antifungal mechanism of ε-poly-L-lysine (ε-PL) against Candida albicans. Methods The standard strains of C. albicans (ATCC 64548 and ATCC 64550) and 50 strains of clinically collected strains were used as experimental strains. Microbroth dilution was performed to determined the minimal inhibitory concentration (MIC), the minimum fungal concentration (MFC), the sessile minimal inhibitory concentration (SMIC50) of ε-PL according to the CLISI-M27 document. The time-growth curve of plankton and biofilm inhibition-time curve of C. albicans was evaluated after the treatment with ε-PL for 48h. The rate of germ tube formation and the germ tube length were observed and recorded within 4h. Fluorescence microscope was used to detect reactive oxygen species (ROS) and spectrophotometry was performed to measure malondialdehyde of C. albicans after the drug treatment. Results The MIC and MFC of ε-PL to C. albicans were 512μg/mL and 1,024μg/mL, respectively. The SMIC50 were the same as the MIC. ε-PL had moderate antifungal effects against C. albicans plankton and biofilm. The fungistatic effect of the membrane became more pronounced with increasing concentrations. The antifungal-time curveresults showed that ε-PL exerted signific antantifungal effects on planktonic bacteria and biofilms of C. albicans at within 12h. The results of the germ tube experiment showed that the high concentration of ε-PL had a significant inhibitory effect on the germ tube formation and germ tube length of C. albicans. The MDA and ROS contents of ε-PL after treating C. albicans showed an upward trend, which was positively correlated with the drug concentration. Conclusion ε-PL had significant antifungal effects on mycoplankton and biofilms of C. albicans. The high concentration of ε-PL can significantly inhibit the main mycelium of C. albicans. The action of ε-PL led to the production of a large amount of ROS in C. albicans and a certain degree of lipid oxidation. Perhaps ε-PL might play a bacteriostatic effect through